SF3A1 - Antibodies - The Human Protein Atlas


	Antibody HPA000690	Antibody HPA030083

ANTIBODY INFORMATION
Provider	Atlas Antibodies Sigma-Aldrich	Atlas Antibodies Sigma-Aldrich
Product name	HPA000690	HPA030083
Host species	Rabbit	Rabbit
Clonalityⁱ The antibodies are designated mAB for monoclonal and pAb for polyclonal.	pAb	pAb
Concentration	0.0566 mg/ml	0.0882 mg/ml
Purity	Affinity purified using the PrEST-antigen as affinity ligand	Affinity purified using the PrEST-antigen as affinity ligand
Released in versionⁱ The release of the Human Protein Atlas in which the antibody was first published.	1.2	6.0
Referencesⁱ References to publications in which the antibody has been used.	1
Proper citation	Atlas Antibodies Cat#HPA000690, RRID:AB_1079933	Atlas Antibodies Cat#HPA030083, RRID:AB_10602183
Validation summaryⁱ All assays through which the antibody has been validated. Assays&annotation provide a detailed description of the different assays. The pie-charts indicate degree of validation.	ICC IHC WB PA	ICC IHC WB PA
IMMUNOCYTOCHEMISTRYⁱ Immunocytochemistry is used to validate the antibody staining and for assessing and validating the protein expression pattern in selected human cell lines. Read more
Validationⁱ Results of validation by standard or enhanced validation. Standard validation is based on concordance with available experimental gene/protein characterization data in the UniProtKB/Swiss-Prot database. Standard validation results in scores Supported, Approved or Uncertain. Enhanced validation is performed using either siRNA knockdown, tagged GFP cell lines or independent antibodies. For the siRNA validation the decrease in antibody-based staining intensity upon target protein downregulation is evaluated. For the GFP validation the signal overlap between the antibody staining and the GFP-tagged protein is evaluated. For the independent antibodies validation the evaluation is based on comparison of the staining of two (or more) independent antibodies directed towards independent epitopes on the protein. For all cases except the siRNA validation, an image representative of the antibody staining pattern is shown. For the siRNA validation, a box plot of the results is shown. Read more	Enhanced - Independent antibodiesⁱ This method is based on comparing the staining pattern using two independent antibodies with no overlapping epitopes. A similar staining pattern (main and additional locations) results in an enhanced antibody validation. Read more Antibody staining overlaps with antibody HPA030083. Immunofluorescent staining of human cell line U-251MG shows localization to nucleoplasm & nuclear speckles. Protein location across all samples similar between independent antibodies	Enhanced - Independent antibodiesⁱ This method is based on comparing the staining pattern using two independent antibodies with no overlapping epitopes. A similar staining pattern (main and additional locations) results in an enhanced antibody validation. Read more Antibody staining overlaps with antibody HPA000690. Immunofluorescent staining of human cell line U2OS shows localization to nucleoplasm & nuclear speckles. Protein location across all samples similar between independent antibodies

Antibody dilution	Human assay: A-431 fixed with PFA, dilution: 1:30 Human assay: U-251MG fixed with PFA, dilution: 1:30 Human assay: U2OS fixed with PFA, dilution: 1:30	Human assay: A-431 fixed with PFA, dilution: 1:50 Human assay: U-251MG fixed with PFA, dilution: 1:50 Human assay: U2OS fixed with PFA, dilution: 1:50
IMMUNOHISTOCHEMISTRYⁱ Immunohistochemistry is used for validating antibody reliability by assessing staining pattern in 44 normal tissues. Validation scores include Enhanced, Supported, Approved and Uncertain. Read more
Validationⁱ Results of validation by standard or enhanced validation based on assessment of antibody performance in 44 normal tissues. Standard validation results in scores Supported, Approved or Uncertain. An image representative of the antibody staining pattern is shown. Enhanced validation results in the score Enhanced and includes two methods: Orthogonal validation and Independent antibody validation. For orthogonal validation, representative images of high and low expression are shown. For independent antibody validation, four images of each independent antibody are displayed. Read more	Enhanced - Independent antibodies Protein distribution across 45 tissues similar between the independent antibodies HPA000690 and HPA030083. Cerebral cortex Colon Kidney Testis	Enhanced - Independent antibodies Protein distribution across 45 tissues similar between the independent antibodies HPA000690 and HPA030083. Cerebral cortex Colon Kidney Testis

Retrievalⁱ Antigen retrieval is a method used to restore/retrieve the epitope (antibody bidning region) of the target protein, cross-linked, and thus masked, during tissue preserving fixative treatment of the tissues. Read more	HIER pH6	HIER pH6
Antibody dilution	1:200	1:1500
Literature conformityⁱ Conformance of the expression pattern with available gene/protein characterization data in scientific literature and data from bioinformatic predictions. UniProt is used as the main source of gene/protein characterization data and when relevant, available publications and other sources of information are researched in depth. Extensive or sufficient gene/protein data requires that there is evidence of existence on a protein level and that a substantial quantity of published experimental data is available from literature and public databases. Limited protein/gene characterization data does not require evidence of existence on a protein level and refers to genes for which only bioinformatic predictions and scarce published experimental data is available.	Consistent with extensive gene/protein characterization data.	Consistent with extensive gene/protein characterization data.
RNA consistencyⁱ Consistency between immunohistochemistry data and consensus RNA levels is divided into five different categories: i) High consistency, ii) Medium consistency, iii) Low consistency, iv) Very low consistency, and v) Cannot be evaluated.	High consistency between antibody staining and RNA expression data.	High consistency between antibody staining and RNA expression data.
WESTERN BLOTⁱ A Western blot analysis is performed on a panel of human tissues and cell lines to evaluate antibody specificity. For antibodies with unreliable result a revalidation using an over-expression lysate is performed. Read more
Validationⁱ Western Blot is used for quality control of the polyclonal antibodies generated in the project. After purification, the antibodies are used to detect bands in a setup of lysate and different tissues. The result is then scored Enhanced, Supported, Approved, or Uncertain. Enhanced validation includes five different methods: Genetic validation, Recombinant expression validation, Independent antibody validation, Orthogonal validation and Capture MS validation. Read more	Enhanced - Capture MSⁱ This method is based on comparison between the molecular weight of the stained band visualized by the antibody against the protein size obtained by a capture MS method in which multiple gel slices are cut out from the electrophoretic separation of cell lysates of RT4 and U-251 and analysed separately by proteomics. The band detected by the antibody should be equivalent to the same of the intended target protein and its peptide(s). Antibody band pattern is confirmed by capture-MS. 230 130 95 72 56 36 28 17 11	Enhanced - Capture MSⁱ This method is based on comparison between the molecular weight of the stained band visualized by the antibody against the protein size obtained by a capture MS method in which multiple gel slices are cut out from the electrophoretic separation of cell lysates of RT4 and U-251 and analysed separately by proteomics. The band detected by the antibody should be equivalent to the same of the intended target protein and its peptide(s). Antibody band pattern is confirmed by capture-MS. 230 130 95 72 56 36 28 17 11 Enhanced - Geneticⁱ This method is based on the knock-down in a suitable cell line of the target protein using genetic methods, in this case siRNA. The staining of the antibody is evaluated by Western blot through analyses of samples from cell lysates before and after knock-down of the corresponding target gene. The results show no or weaker band in the lysate from the knock-down cell line. Downregulation visible in both siRNA lanes Lane 1: Marker [kDa] 250, 130, 95, 72, 55, 36, 28, 17, 10 Lane 2: siRNA 1 (U2OS) Lane 3: siRNA 2 (U2OS) Lane 4: Scrambled (U2OS) Target mass (kDa): 88.9 Total protein image

Antibody dilution	1:250	Capture MS: 1:250 Genetic: 1:98
PROTEIN ARRAY
Validationⁱ A protein array containing 384 different antigens including the antibody target is used to analyse antibody specificity. Depending on the array interaction profile the antibody is scored as Supported, Approved, or Uncertain. Read more	Supported Pass with single peak corresponding to interaction only with its own antigen. Antibody specificity analysis with protein arrays. Predicted and matching interactions are shown in green.	Supported Pass with single peak corresponding to interaction only with its own antigen. Antibody specificity analysis with protein arrays. Predicted and matching interactions are shown in green.

Antibody dilution	1:1000	1:3000
RELEVANT PUBLICATIONS
	The nuclear splicing factor RNA binding motif 5 promotes caspase activation in human neuronal cells, and increases after traumatic brain injury in mice Jackson TC et al J Cereb Blood Flow Metab 2015;35(4):655-66 Application: WB
ANTIGEN INFORMATION
Antigen	Recombinant protein fragment	Recombinant protein fragment
Length (aa)	107	175
Antigen sequence	`TEDSLMPEEEFLRRNKGPVSIKVQVPNMQDKTEWKLNGQVLVFTLPLTDQ VSVIKVKIHEATGMPAGKQKLQYEGIFIKDSNSLAYYNMANGAVIHLALK ESGGRKK`	`YPPPEVRNIVDKTASFVARNGPEFEARIRQNEINNPKFNFLNPNDPYHAY YRHKVSEFKEGKAQEPSAAIPKVMQQQQQTTQQQLPQKVQAQVIQETIVP KEPPPEFEFIADPPSISAFDLDVVKLTAQFVARNGRQFLTQLMQKEQRNY QFDFLRPQHSLFNYFTKLVEQYTKI`
Matching transcripts	SF3A1-201 - ENSP00000215793 [99%]	SF3A1-201 - ENSP00000215793 [100%]
Matching mouse transcripts	ENSMUSP00000002198 [97%] ENSMUSP00000057293 [22%] ENSMUSP00000010520 [22%]	ENSMUSP00000002198 [99%] ENSMUSP00000011450 [27%] ENSMUSP00000078469 [23%]
ANTIGEN VIEWⁱ The protein browser displays the antigen location on the target protein(s) and the features of the target protein. The tabs at the top of the protein view section can be used to switch between the different splice variants to which an antigen has been mapped. At the top of the view, the position of the antigen (identified by the corresponding HPA identifier) is shown as a green bar. A yellow triangle on the bar indicates a <100% sequence identity to the protein target. Below the antigens, the maximum percent sequence identity of the protein to all other proteins from other human genes is displayed, using a sliding window of 10 aa residues (HsID 10) or 50 aa residues (HsID 50). The region with the lowest possible identity is always selected for antigen design, with a maximum identity of 60% allowed for designing a single-target antigen (read more). The curve in blue displays the predicted antigenicity i.e. the tendency for different regions of the protein to generate an immune response, with peak regions being predicted to be more antigenic.The curve shows average values based on a sliding window approach using an in-house propensity scale. (read more). If a signal peptide is predicted by a majority of the signal peptide predictors SPOCTOPUS, SignalP 4.0, and Phobius (turquoise) and/or transmembrane regions (orange) are predicted by MDM, these are displayed. Low complexity regions are shown in yellow and InterPro regions in green. Common (purple) and unique (grey) regions between different splice variants of the gene are also displayed (read more), and at the bottom of the protein view is the protein scale.
SF3A1-201